A single amino acid exchange inverts susceptibility of related receptor tyrosine kinases for the ATP-site inhibitor STI-571

Abstract

The tyrosine kinase inhibitor STI-571 potently blocks BCR-Abl, platelet-derived growth factor (PDGF) a- and b-receptors, and c-Kit kinase activity. However Flt3, a receptor tyrosine kinase closely related to PDGF receptors and c-Kit, is not inhibited by STI-571. Sequence alignments of different kinases and indications from the crystal structure of the STI-571 Abl kinase complex revealed amino acid residues that are probably crucial for this activity profile. It was predicted that Flt3 Phe-691 in the b5 strand may sterically prevent interaction with STI-571. The point mutants Flt3 F691T and PDGFb-receptor T681F were constructed, and kinase assays showed that the Flt3 mutant but not the PDGFb-receptor mutant is inhibited by STI-571. Docking of STI-571 into computer models of the PDGFb-receptor and Flt3 kinase domains and comparison with the crystal structure of the STI-571 Abl kinase complex indicated very similar binding sites among the three nonphosphorylated kinases, suggesting corresponding courses of their Asp-Phe-Gly motifs and activation loops. Accordingly, we obsd. reduced sensitivity of preactivated compared with nonactivated PDGFR-b for the inhibition by STI-571. Courses of the activation loop that collide with STI-571 binding explain its inactivity toward other kinases such as the insulin receptor. The binding site models of PDGFR-b and Flt3 were applied to predict structural approaches for more selective PDGFb-receptor inhibitors.